人类青光眼和青光眼动物模型中神经胶质细胞的改变

青光眼是主要的致盲眼病之一,其特征性表现为视乳头的凹陷性萎缩和视野的特征性缩小.青光眼的病因复杂,但却有共同的病理结局即视网膜神经节细胞的进行性凋亡.因此对青光眼的研究多集中在视网膜神经节细胞的改变及其机制上.但近年来的研究结果表明,作为神经系统另一大类细胞即神经胶质细胞在青光眼的病理过程中也发生了活化,并且其活化与视网膜神经节细胞的改变及疾病的发生、发展密切相关.所以对青光眼中神经胶质细胞的研究也越来越深入.笔者就目前对神经胶质细胞在人类青光眼及青光眼动物模型中的改变进行综述,以供同道参考.     

胶质细胞活化相关炎症在青光眼视神经病变中的研究进展 △

青光眼是一种以视网膜神经节细胞(RGCs)及其轴突凋亡为特征的视神经退行性病变。越来越多的研究表明炎症和免疫反应在青光眼视神经病变中具有重要作用。在高眼压动物实验中,抑制早期胶质细胞活化及减少炎症因子释放,对RGCs和视神经具有保护作用。本文重点就星形胶质细胞和小胶质细胞活化及其产生的炎症因子,特别是视网膜及视盘部胶质细胞释放的炎症因子在青光眼视神经病变过程中的作用及相关机制进行综述,希望能给青光眼的研究和治疗带来新的启发。     

神经生长因子-β、脑源性神经营养因子、胶质细胞源性神经营养因子对体外压力作用下视网膜神经细胞的保护作用

高眼压是青光眼最主要的致病因素，青光眼的视野丧失是节细胞死亡的结果，供应视网膜的神经营养因子被剥夺、细胞内离子的失衡等参与了细胞的损害过程。神经生长因子(NGF)、脑源性神经营养因子(BDNF)、胶质细胞源性神经营养因子(GDNF)、睫状节神经营养因子(CNTF)等可减少实验性高眼压缺血、视神经切断后的视网膜神经节细胞的死亡。为     

米诺环素对视网膜神经节细胞保护作用的研究进展

视网膜神经节细胞作为中枢神经系统的一部分,是青光眼和视网膜疾病的主要受损细胞。米诺环素是一种半合成的四环素类衍生物,除广谱抗菌功效外,还具有抗氧化、抗凋亡和抑制小胶质细胞活化的作用,对神经元具有一定的保护作用。研究证明米诺环素对视网膜神经节细胞也具有保护作用,在视神经外伤、青光眼和多种视网膜疾病的研究中均显示了不同程度的效果。本文就米诺环素对视网膜神经节细胞的神经保护作用及其作用机制作一综述。     

慢性高眼压大鼠视网膜神经胶质细胞组织病理学改变

目的 研究青光眼视网膜神经胶质细胞组织病理学改变及其在青光眼视网膜神经节细胞损伤中的作用机制.方法 对照实验研究.选用造模成功的慢性高眼压雄性SD大鼠72只,眼压＞22 mm Hg(1 mm Hg=0.133 kPa).右眼为慢性高眼压眼,左眼为假手术对照眼.根据慢性高眼压模型建立的时间(手术结束时开始计算),将实验鼠分为6组(2、12 h,1 d,1、4、8周),每组12只鼠.正常组SD大鼠12只,平均眼压12.56 mm Hg.分别取实验各组(慢性高眼压)、假手术组及正常组大鼠4只眼球,冰冻切片行胶质纤维酸性蛋白(GFAP)免疫组织化学染色,在激光共焦显微镜下观察视网膜星形胶质细胞及Müller细胞的GFAP表达情况;分别取实验各组(慢性高眼压)和正常组大鼠4只眼球,在视网膜铺片上进行GFAP免疫组织化学染色,进行星形胶质细胞计数并观察其形态;分别取实验各组(慢性高眼压)、假手术组及正常大鼠4只眼球的鼻侧半视网膜,在视网膜铺片上行小胶质细胞标记物OX42免疫组织化学染色,进行小胶质细胞计数并观察其形态;取剩余的颢侧中周部视网膜,半薄切片行甲苯胺蓝染色并进行Müller细胞计数.对不同时间点慢性高眼压组与正常组大鼠细胞表达数最进行比较,采用单因素多水平设计定量资料的方差分析.结果 慢性高眼压模型建立后2 h,即有活化的小胶质细胞出现;1 d后小胶质细胞的数量开始增加,为(327.40±68.32)个/mm2;1周后小胶质细胞的数量达到高峰,为(965.06±86.63)个/mmw,与正常组小胶质细胞数最比较,差异有统计学意义(F=196.56,P＜0.01);其后小胶质细胞数量逐渐减少.慢性高眼压模型建立后12 h,星形胶质细胞及Müller细胞开始呈现活化状态;4周时两种细胞的活化程度达到高峰,以后活化程度逐渐下降,且活化的星形胶质细胞在结构上出现明显异常,表现为星形胶质细胞突起变得粗短、僵硬,胞体的星型结构破坏;但慢性高眼压组视网膜星形胶质细胞及Müller细胞数量与正常组相比,差异均无统计学意义(F=1.36,1.89;均P＞0.05).结论 在慢性高眼压条件下,小胶质细胞可能是视网膜最早发生病理学改变的组织;活化的星形胶质细胞可出现明显的形态和结构变化,其结果不仅将加速神经节细胞的损伤,同时也会形成不利于神经节细胞轴突再生的视网膜微环境.     

小胶质细胞极化在青光眼视神经损伤发病机制及治疗中的研究进展

青光眼是多因素介导的以视网膜神经节细胞凋亡、视神经萎缩和视野缺损为特征的神经退行性眼病,发病机制尚不明确.小胶质细胞是视网膜内常驻的免疫细胞,它可分为经典激活M1型和替代激活M2型,随着眼压改变以及视网膜神经节细胞损伤修复过程的进展,小胶质细胞的极性呈现动态变化,可产生多种具有神经毒性或神经保护作用的细胞因子.近年来,...     

青光眼视网膜神经节细胞的凋亡

青光眼是一种主要致盲性眼病,该文就视网膜神经节细胞是怎样死亡的,为什么该细胞死亡程序被激发,以及未来可有效地治疗青光眼的可能性这几方面进行综述。     

视网膜神经胶质细胞与青光眼

青光眼视神经病变以进行性的视网膜神经节细胞丧失和相应的视功能损害为特征.导致青光眼视神经病变的病理生理机制尚不明确.在中枢神经系统,神经胶质细胞参与了神经元在受到机械、缺血、缺氧等损伤后的病理变化过程.胶质细胞参与青光眼视神经病变的机制复杂多样,但部分机制与中枢神经系统相似.本文就视网膜神经胶质细胞的生理功能、动物青光眼模型建立后胶质细胞的反应以及针对胶质细胞的青光眼视神经保护治疗手段做一综述.     

藏红花提取液对兔慢性高眼压模型中视网膜相关生长蛋白表达的影响

青光眼患者由于慢性高眼压引起眼部血液循环异常、视神经轴浆流受阻[1,2],加之谷氨酸浓度的升高,反应性胶质细胞的活化及一氧化氮的毒性作用[3],导致视网膜神经节细胞(RGC)凋亡和视神经的损伤,最终导致失明.     

青光眼视网膜神经节细胞的凋亡

青光眼是一种主要致盲性眼病，该文就视网膜神经节细胞是怎样死亡的，为什么该细胞死亡程序被激发，以及未来可有效地治疗青光眼的可能性这几方面进行综述。     

视网膜神经节细胞无血清上清培养液诱导大鼠视网膜干细胞的分化

目的　探讨视网膜神经节细胞无血清上清培养液对视网膜干细胞分化的影响。方法　分离大鼠视网膜干细胞和视网膜神经节细胞;采用免疫荧光法鉴定体外培养的大鼠视网膜干细胞与视网膜神经节细胞,视网膜干细胞以Nestin抗体进行鉴定,视网膜神经节细胞以Thy-1抗体进行鉴定;以视网膜神经节细胞无血清上清培养液培养视网膜干细胞,以不加入条件培养液培养的视网膜干细胞为对照组,收集分化细胞,采用qPCR法检测Nestin、Pax6、Thy-1及Brn-3的基因表达。结果　培养的视网膜干细胞Nestin抗体染色阳性,视网膜神经节细胞Thy-1抗体染色阳性。培养视网膜干细胞72 h后,与对照组相比,无血清上清培养液组细胞Nestin和PAX6基因相对表达量降低,差异均有统计学意义（均为P<0.000 1）;Thy-1和Brn-3基因相对表达量升高,差异均有统计学意义（均为P<0.05）。结论　视网膜神经节细胞无血清上清培养液能够诱导视网膜干细胞分化为视网膜神经节样细胞。     

Houttuynia cordata Thunb rescues retinal ganglion cells through inhibiting microglia activation in a rat model of retinal ischemia-reperfusion

AIM: To observe the melanin change of the retinal pigment epithelium (RPE) and choroid in the convalescent stage of Vogt-Koyanagi-Harada (VKH). METHODS: A retrospective study was performed on 40 eyes of 20 patients in the convalescent stage of VKH. Fundus photography (FP), multispectral imaging (MSI), and optical coherence tomography (OCT) were performed. RESULTS: In the VKH convalescent stage, focal RPE melanin accumulation (FRMA) was detected in 34 eyes (85%) on MSI and in 7 eyes (17.5%) on FP. FRMA was limited to the previous retinal detachment area in all 28 eyes (FRMA was detected in 34 eyes on MSI, which were enrolled, and 6 eyes lacked data in the acute stage). Sunset-glow fundus was detected in 20 eyes (50%) on FP. The mean density of FRMA in a 1-mm-diameter circular area of the fovea was 0.04±0.07 on MSI, which was significantly correlated with sunset-glow fundus (ρ=0.467, P=0.02). CONCLUSION: In the VKH convalescent stage, FRMA is derived from the RPE melanin change, and sunset-glow fundus is derived from the choroid melanin change. A higher density of FRMA in the fovea and sunset-glow fundus represents more serious depigmentation of melanin.     

Increased activity of cyclooxygenase-2 signals early neurodegenerative events in the rat retina following transient ischemia

Following transient retinal ischemia, there is neuronal cell death, breakdown of the blood-retinal barrier, activation of microglia and infiltration by hematogenous cells. The early appearance of cyclooxygenase-2 (COX-2) following an ischemic event may be responsible for signaling some of the responses that lead to neurodegeneration. We have determined the time courses of changes in protein levels and cellular localizations of COX-2 in the rat retina after transient ischemia. In the normal rat retina, COX-2 immunoreactivity was present in neurons in the INL and ganglion cell layer (GCL). Six to 12 hr after ischemia, COX-2 immunoreactivity was upregulated/induced in horizontal cells, amacrine cells, retinal ganglion cells, displaced amacrine cells of the INL and GCL, and Müller cells. The NMDA-receptor antagonist, MK801, blocked the increased COX-2 protein level and COX-2 immunoreactivity in neurons of the INL and GCL, but did not affect the induction of COX-2 in Müller cells after ischemia. The selective COX-2 inhibitor, SC-58236, prevented apoptotic cell death and was neuroprotective against loss of retinal ganglion cells after ischemia. Following transient ischemia, the selective COX-2 inhibitor did not prevent breakdown of the blood-retinal barrier or activation of microglia. However, the selective COX-2 inhibitor reduced infiltration of hematogenous cells into the retina. These results suggest that the early, increased activity of COX-2 signals neurodegenerative events following retinal ischemia.     

Involvement of glycinergic neurons in the diminished surround activity of ganglion cells in the dark-adapted rabbit retina.

Previous studies have reported that the surround responses of retinal ganglion cells weaken or disappear upon dark adaptation. The mechanism(s) by which this occurs is largely unknown, although changes in activity of retinal dopaminergic neurons have been implicated. In the light-adapted rabbit retina, the surround ON responses of OFF-center ganglion cells have been shown to be markedly reduced or abolished by a dopamine antagonist. This effect of a dopamine antagonist was recently shown to be reversed by the glycine antagonist strychnine and by compounds that elevate intracellular cAMP levels. The present study was conducted to determine whether strychnine and cAMP-elevating compounds could bring out the surround ON responses in OFF-center ganglion cells that are diminished upon dark adaptation. Extracellular recordings of OFF-center brisk ganglion cells were made from isolated, superfused retinal preparations. During the course of dark adaptation, the surround ON responses of many cells decreased markedly. Application of low micromolar concentrations of strychnine to the bathing solution brought out the surround ON responses in both brisk-transient and brisk-sustained OFF-center ganglion cells. The center OFF responses of these cells, on the other hand, were not enhanced by strychnine. Of the cAMP-elevating compounds tested, 8-(4-chlorophenylthio) cyclic AMP was the most effective in bringing out the surround ON responses in dark-adapted OFF-center ganglion cells. The effects of bath application of this cAMP analog were very similar to those of strychnine. The findings from this study suggest that under dark-adapted conditions glycinergic neurons inhibit the surround component of OFF-center ganglion cells. The release of glycine from these neurons is suggested to be regulated by a cAMP-dependent mechanism.     

Detection of early neuron degeneration and accompanying microglial responses in the retina of a rat model of glaucoma

PURPOSE: To characterize the early reaction of retinal ganglion cells (RGCs) in a rat model of glaucoma using in vivo imaging and to examine the involvement of retinal microglia in glaucomatous neuropathy. METHODS: Glaucoma was induced in adult female Sprague-Dawley rats by cauterizing two episcleral veins, which resulted in a 1.6-fold increase in intraocular pressure (IOP). Retinal ganglion cells were retrogradely labeled with the fluorescent dye, 4-[didecylaminostyryl]-N-methyl-pyridinium-iodide (4-Di-10ASP) and monitored in vivo after elevation of IOP using fluorescence microscopy imaging. The number of RGCs was quantified on retinal flatmounts. Dying RGCs were surrounded by activated microglia that became visible after taking up the fluorescent debris. Immunocytochemistry was conducted to characterize further the ganglion cells and microglia. RESULTS: Cauterizing two of the four episcleral veins resulted in a consistent increase of IOP to 25.3 +/- 2.0 mm Hg, as measured with a handheld tonometer. IOP remained high for at least 3 months in glaucomatous eyes. The earliest sign of RGC death was detected in anesthetized animals 20 hours after induction of glaucoma. RGCs continued to decrease in number over time, with 40% of RGCs having degenerated after 2.5 months. Fundoscopic examination of the optic nerve head revealed cupping 2 months after induction of glaucoma. In addition, microglia were detected on retinal flatmounts as early as 72 hours after induction. Activated microglia and RGCs were also identified immunocytochemically, with an antibody against ionized calcium-binding adaptor molecule (Iba)-1 and an antibody specific to the 200-kDa subunit of the neurofilament protein, respectively. CONCLUSIONS: Occlusion of episcleral veins is a reproducible method that mimics human glaucoma, with chronically elevated IOP-induced RGC loss. This study shows that in vivo imaging permits the detection of ganglion cells in the living animal in the early stages of the disease and highlights the importance of in vivo imaging in understanding ophthalmic disorders such as glaucoma. Secondly, activation of intraretinal microglia coincides with degeneration of RGCs in glaucoma.     

Hydrogen-rich saline protects retina against glutamate-induced excitotoxic injury in guinea pig

Molecular hydrogen (H(2)) is an efficient antioxidant that can selectively reduce hydroxyl radicals and inhibit oxidative stress-induced injuries. We investigated the protective effects and mechanism of hydrogen-rich saline in a glutamate-induced retinal injury model. Retinal excitotoxicity was induced in healthy guinea pigs by injecting glutamate into the vitreous cavity. After 30?min, hydrogen-rich saline was injected into the vitreous cavity, the peritoneal cavity or both. Seven days later, the retinal stress response was evaluated by examining the stress biomarkers, inducible nitric-oxide synthase (iNOS) and glucose-regulated protein 78 (GRP78). The impaired glutamate uptake was assessed by the expression of the excitatory amino acid transporter 1(EAAT-1). The retinal histopathological changes were investigated, focusing on the thicknesses of the entire retina and its inner layer, the number of cells in the retinal ganglion cell layer (GCL) and the ultrastructure of the retinal ganglion cells (RGCs) and glial cells. Compared with the glutamate-induced injury group, the hydrogen-rich saline treatment reduced the loss of cells in the GCL and thinning of the retina and attenuated cellular morphological damage. These improvements were greatest in animals that received H(2) injections into both the vitreous and the peritoneal cavities. The hydrogen-rich saline also inhibited the expression of glial fibrillary acidic protein (GFAP) in Müller cells, CD11b in microglia, and iNOS and GRP78 in glial cells. Moreover, the hydrogen-rich saline increased the expression of EAAT-1. In conclusion, the administration of hydrogen-rich saline through the intravitreal or/and intraperitoneal routes could reduce the retinal excitotoxic injury and promote retinal recovery. This result likely occurs by inhibiting the activation of glial cells, decreasing the production of the iNOS and GRP78 and promoting glutamate clearance.     

嘌呤P2X7受体在视网膜变性类疾病中的作用研究进展

视网膜变性类疾病是世界上主要的一类致盲性眼病,感光细胞等视网膜神经元细胞损伤及凋亡是其共同的病理基础.嘌呤能离子通道型7(P2 X7)受体在视网膜多种类型的细胞均有表达.研究发现,P2 X7受体激活及其介导的信号异常表达与视网膜神经元细胞变性死亡过程相关,而阻断或下调P2 X7受体的激活表达可显著减少视网膜胶质细胞激活...     

Involvement of the CD200 receptor complex in microglia activation in experimental glaucoma

The interaction of the myeloid restricted molecule CD200R with its widely expressed ligand CD200 is involved in the down-regulation of microglia activation. In the present study, we examined the involvement of CD200R in microglia activation in experimental ocular hypertension to determine the role of microglia activation in retinal ganglion cell (RGC) death, the key pathological event in glaucoma. Experimental glaucoma was induced in adult Brown Norway rats by sclerosis of the episcleral veins with the injection of hypertonic saline. Immunohistochemical methods were used to determine the involvement of microglia using GFAP, CD45, OX42 and OX41 and the involvement of CD200 and CD200R in the optic nerve head. Our data demonstrate the increased presence of microglia within the optic nerve head during ocular hypertension, identified by positive staining with OX42 and OX41. The peak of microglia correlates with peak in RGC death at days 20-27 (T3) post OHT induction. In addition, CD200 and CD200R positive cells were increased in ocular hypertensive eyes. Increased expression of CD200 was detected in the early phase (days 1-7; T1) of OHT and decreased over time, whilst the expression of CD200R was detected in the middle phase (days 20–27; T3) of OHT, correlating with the increase in microglia markers. Changes in the expression of CD200R/CD200 occur early in experimental glaucoma and precede the peak in microglia infiltration and RGC death, suggesting that CD200R-positive microglia play an important role in the initiation of RGC death during OHT, indicating a potential area for therapeutic intervention in treating glaucoma.     

视网膜缺血-再灌注损伤中小胶质细胞对微循环的破坏作用

目的　探讨视网膜缺血-再灌注损伤(retinal ischemia-reperfusion injury,RIRI)中小胶质细胞活化与视网膜微循环损伤的关系及作用机制。方法　160只雄性C57BL/6小鼠右眼均采用前房灌注建立RIRI模型为RIRI组，左眼不作处理为正常对照组。在损伤后24 h、48 h、72 h分别进行视网膜冰冻切片、视网膜铺片免疫荧光染色检测小胶质细胞的活化情况，检测相关缺氧因子及炎症因子的表达，与正常对照组比较，研究小胶质细胞的激活状态与微循环损伤的关系，并初步分析其作用机制。结果　视网膜微循环结构损伤观察结果显示，与正常对照组相比，RIRI后24 h组大部分血管仍呈正常形态；RIRI后48 h组，闭塞的血管数量增多；RIRI后72 h组，血管损伤明显加重。浅层毛细血管密度正常对照组，RIRI后24 h、48 h、72 h组间两两比较差异均无统计学意义（均为P>0.05）。而深层血管网毛细血管密度RIRI后72 h组与其余3组相比明显减少，差异均有统计学意义（均为P<0.05），其余3组间差异均无统计学意义（均为P>0.05）。视网膜冰冻切片检测显示，RIRI后24 h组与正常对照组各层视网膜中抗离子钙接头蛋白（ionized calcium bindingadaptor molecule-1，Iba-1）阳性细胞数差异无统计学意义（P>0.05）。RIRI后48 h组各层中Iba-1阳性细胞数与正常对照组及24 h组差异均有统计学意义（均为P<0.05）。RIRI后72 h组各层中Iba-1阳性细胞数与正常对照组、24 h组、48 h组差异均有统计学意义（均为P<0.05）。视网膜铺片结果显示，RIRI后48 h组和72 h组活化的小胶质细胞数明显增加，与正常对照组和RIRI后24 h组在两个层次毛细血管中差异均有统计学意义（均为P<0.05），而72 h组小胶质细胞的活化达到高峰值，与其余组差异均有统计学意义（均为P<0.05）。RT-PCR检测结果显示:血管内皮生长因子和缺氧诱导因子-1α的缺血缺氧因子，肿瘤坏死因子-α和白细胞介素-1β的炎症因子在正常对照组与RIRI后24 h组、48 h组、72 h组，组间比较差异均有统计学意义（均为P<0.05）。结论　激活的小胶质细胞在RIRI中发挥了对微循环的破坏作用，损伤早期抑制小胶质细胞的活化可能成为此类疾病治疗的新思路。     

Evoked expression of the glutamate transporter GLT-1c in retinal ganglion cells in human glaucoma and in a rat model

PURPOSE: Glaucoma is a common disease of the eye, a key characteristic consequence of which is the death of retinal ganglion cells. The cause of this loss is unknown, though glutamate-mediated toxicity has been implicated. Glutamate transporters are key regulators of glutamate; therefore, the purpose of the study was to determine whether unusual excitation is associated with unusual expression of one or more transporters. METHODS: The expression of a splice variant of the glutamate transporter GLT-1 (EAAT2) was examined in normal and glaucomatous retinas from humans and rats. RESULTS: In normal eyes of humans and rats, GLT-1c was expressed only in photoreceptors. In glaucoma, there was additional robust expression of GLT-1c in retinal ganglion cells, including occasional displaced ganglion cells. Conversely, cells such as displaced amacrine cells and amacrine cells were unlabeled. CONCLUSIONS: The induction of GLT-1c expression by retinal ganglion cells supports the notion that an anomaly or anomalies in glutamate homeostasis may be evident in glaucoma and that such anomalies selectively influence retinal ganglion cells. By analogy to in vitro experiments in which elevated glutamate levels induce expression of glutamate transporters, the authors hypothesize that expression of GLT-1c may represent an attempt by retinal ganglion cells to protect themselves against elevated levels of glutamate. Such anomalies in glutamate levels cannot be restricted to the ganglion cell layer, as this would not have affected displaced ganglion cells. GLT-1c may be a useful indicator of the extent of stress of the retinal ganglion cells and thus a tool for examining outcomes of potential therapeutic and experimental interventions.